Severe Neonatal Opioid Withdrawal Requiring Pharmacotherapy: Impact of Region of Residence.

OBJECTIVES: Our objective was to evaluate the trend and to assess the impact of maternal region of residence in Western New York (WNY), on severe neonatal opioid withdrawal syndrome (NOWS). STUDY DESIGN: Term infants' born at gestational age greater than or equal to 37 weeks with severe NOWS, defined as withdrawal resulting in the receipt of pharmacologic therapy from WNY admitted to our neonatal intensive care unit (NICU) from January 1, 2008 to December 31, 2016, were included. Severe NOWS admissions to our NICU from the following five regions were controlled with birth and insurance data: (1) Urban North, (2) Erie Coastal, (3) Niagara Frontier, (4) Southern Tier, and (5) Urban South. RESULTS: "Urban South" residence was associated with an increased risk of severe NOWS (adjusted odds ratio = 1.8, 97.5% confidence interval: 1.1-2.9). The trend in admission for severe NOWS doubled between 2008 to 2010 and 2014 to 2016 (p = 0.01). More infants born to maternal nonprescribed opioid users were placed in foster care at discharge (36.5 vs. 1.9%, p < 0.001). CONCLUSION: In WNY, neonates born to mothers from the "Urban South" were twice at risk of being admitted for severe NOWS. One-third of infants with severe NOWS after nonprescribed opioid use were placed in foster care. Implementing targeted strategies at the community level may help improve outcomes in NOWS. KEY POINTS: · Maternal region of residence is a risk factor for severe neonatal opioid withdrawal.. · Admissions for severe neonatal opioid withdrawal trended up from 2008 to 2010 to 2014 to 2016.. · One-third of the infants born to mothers on nonprescribed opioids were discharged to foster care..

Hydrocortisone to Improve Survival without Bronchopulmonary Dysplasia.

BACKGROUND: Bronchopulmonary dysplasia is a prevalent complication after extremely preterm birth. Inflammation with mechanical ventilation may contribute to its development. Whether hydrocortisone treatment after the second postnatal week can improve survival without bronchopulmonary dysplasia and without adverse neurodevelopmental effects is unknown. METHODS: We conducted a trial involving infants who had a gestational age of less than 30 weeks and who had been intubated for at least 7 days at 14 to 28 days. Infants were randomly assigned to receive either hydrocortisone (4 mg per kilogram of body weight per day tapered over a period of 10 days) or placebo. Mandatory extubation thresholds were specified. The primary efficacy outcome was survival without moderate or severe bronchopulmonary dysplasia at 36 weeks of postmenstrual age, and the primary safety outcome was survival without moderate or severe neurodevelopmental impairment at 22 to 26 months of corrected age. RESULTS: We enrolled 800 infants (mean [±SD] birth weight, 715±167 g; mean gestational age, 24.9±1.5 weeks). Survival without moderate or severe bronchopulmonary dysplasia at 36 weeks occurred in 66 of 398 infants (16.6%) in the hydrocortisone group and in 53 of 402 (13.2%) in the placebo group (adjusted rate ratio, 1.27; 95% confidence interval [CI], 0.93 to 1.74). Two-year outcomes were known for 91.0% of the infants. Survival without moderate or severe neurodevelopmental impairment occurred in 132 of 358 infants (36.9%) in the hydrocortisone group and in 134 of 359 (37.3%) in the placebo group (adjusted rate ratio, 0.98; 95% CI, 0.81 to 1.18). Hypertension that was treated with medication occurred more frequently with hydrocortisone than with placebo (4.3% vs. 1.0%). Other adverse events were similar in the two groups. CONCLUSIONS: In this trial involving preterm infants, hydrocortisone treatment starting on postnatal day 14 to 28 did not result in substantially higher survival without moderate or severe bronchopulmonary dysplasia than placebo. Survival without moderate or severe neurodevelopmental impairment did not differ substantially between the two groups. (Funded by the National Institutes of Health; ClinicalTrials.gov number, NCT01353313.).

Engineering encodable lanthanide-binding tags into loop regions of proteins.

Lanthanide-binding tags (LBTs) are valuable tools for investigation of protein structure, function, and dynamics by NMR spectroscopy, X-ray crystallography, and luminescence studies. We have inserted LBTs into three different loop positions (denoted L, R, and S) of the model protein interleukin-1ß (IL1ß) and varied the length of the spacer between the LBT and the protein (denoted 1−3). Luminescence studies demonstrate that all nine constructs bind Tb3+ tightly in the low nanomolar range. No significant change in the fusion protein occurs from insertion of the LBT, as shown by two X-ray crystallographic structures of the IL1ß-S1 and IL1ß-L3 constructs and for the remaining constructs by comparing the 1H−15N heteronuclear single-quantum coherence NMR spectra with that of the wild-type IL1ß. Additionally, binding of LBT-loop IL1ß proteins to their native binding partner in vitro remains unaltered. X-ray crystallographic phasing was successful using only the signal from the bound lanthanide. Large residual dipolar couplings (RDCs) could be determined by NMR spectroscopy for all LBT-loop constructs and revealed that the LBT-2 series were rigidly incorporated into the interleukin-1ß structure. The paramagnetic NMR spectra of loop-LBT mutant IL1ß-R2 were assigned and the Δχ tensor components were calculated on the basis of RDCs and pseudocontact shifts. A structural model of the IL1ß-R2 construct was calculated using the paramagnetic restraints. The current data provide support that encodable LBTs serve as versatile biophysical tags when inserted into loop regions of proteins of known structure or predicted via homology modeling.

Lanthanide-binding tags with unnatural amino acids: sensitizing Tb3+ and Eu3+ luminescence at longer wavelengths.

Lanthanide-binding tags (LBTs) are small, genetically encoded, versatile protein fusion partners that selectively bind lanthanide ions with high affinity. The LBT motif features a strategically positioned tryptophan residue that sensitizes Tb3+ luminescence upon excitation at 280 nm. Herein, we describe the preparation of new LBT peptides that incorporate unnatural amino acids in place of tryptophan, and which sensitize both Tb3+ and Eu3+ luminescence at lower energies. We also report the semisynthesis of proteins tagged with these new LBTs using native chemical ligation. This expands the scope of LBTs and will enable their wider use in luminescence applications.

Characterization of a 1:1 Cu-O2 adduct supported by an anilido imine ligand.

Copper(I) complexes of sterically hindered anilido imine ligands o-C6H4{N(C6H3(i)Pr2)}{C(R)=NC6H3(i)Pr2}- (L(1), R = H; L(2), R = CH3) have been prepared and characterized by spectroscopic and X-ray crystallographic methods. These complexes are highly reactive with O2, and in the case of L2 the product of low-temperature oxygenation was fully characterized by spectroscopic, X-ray crystallographic, and computational methods. The resonance Raman spectrum features an isotope-sensitive vibration at 974 cm(-1) (Delta(18O) = 66 cm(-1)), consistent with assignment as an O-O stretch. Despite the asymmetric coordination environment provided by the supporting anilido imine ligand, the X-ray crystal structure confirms rather symmetric side-on binding of the O2 moiety to the copper center, and the O-O bond length of 1.392(2) Angstroms indicates that this intermediate has significant Cu(III)-peroxo character. Theoretical calculations support this interpretation and predict that while a fully optimized end-on singlet geometry can be obtained, it is higher in energy than the side-on isomer by 3.5 kcal mol(-1) at the CASPT2/TZP level.

Reactivity of a 1:1 copper-oxygen complex: isolation of a Cu(II)-o-iminosemiquinonato species.

While a 1:1 Cu-O2 adduct is generally unreactive with organic substrates, phosphines displace O2 via an associative process and added Cu(I) leads to a novel internal ligand oxidation to yield a Cu(II)-o-iminosemiquinone complex.

Mixed metal bis(mu-oxo) complexes with [CuM(mu-O)2]n+(M = Ni(III) or Pd(II)) cores.

Two highly reactive heterodinuclear bis(mu-oxo) complexes were prepared by combining mononuclear peroxo species with reduced metal precursors at -80 degrees C and were identified by UV-vis, EPR/NMR, and resonance Raman spectroscopy, with corroboration in the case of the CuPd system from density functional calculations.

A new class of (mu-eta2:eta2-disulfido)dicopper complexes: synthesis, characterization, and disulfido exchange.

Rare examples of (mu-eta2:eta2-disulfido)dicopper complexes have been prepared from Cu(I) and Cu(II) complexes of beta-diketiminate and anilido-imine supporting ligands. A novel byproduct derived from sulfur functionalization of the methine position of a beta-diketiminate ligand was identified. DFT calculations on [(LCu)2X2] (L = beta-diketiminate, X = O or S) complexes rationalize the absence of a bis(mu-sulfido)dicopper isomer, [Cu2(mu-S)2](2+), in the synthetic reactions, yet predict that a [Cu2(mu-S)2](0) core is a stable product of 2-electron reduction of the [Cu2(mu-eta2:eta2-S2)](2+) unit. Exchange of the disulfido ligand was discovered upon reaction of a (mu-eta2:eta2-disulfido)dicopper complex with a Cu(I) reagent.

Mechanistic studies on the formation and reactivity of dioxygen adducts of diiron complexes supported by sterically hindered carboxylates.

Dioxygen activation by enzymes such as methane monooxygenase, ribonucleotide reductase, and fatty acid desaturases occurs at a nonheme diiron active site supported by two histidines and four carboxylates, typically involving a (peroxo)diiron(III,III) intermediate in an early step of the catalytic cycle. Biomimetic tetracarboxylatodiiron(II,II) complexes with the familiar "paddlewheel" topology comprising sterically bulky o-dixylylbenzoate ligands with pyridine, 1-methylimidazole, or THF at apical sites readily react with O(2) to afford thermally labile peroxo intermediates that can be trapped and characterized spectroscopically at low temperatures (193 K). Cryogenic stopped-flow kinetic analysis of O(2) adduct formation carried out for the three complexes reveals that dioxygen binds to the diiron(II,II) center with concentration dependences and activation parameters indicative of a direct associative pathway. The pyridine and 1-methylimidazole intermediates decay by self-decomposition. However, the THF intermediate decays much faster by oxygen transfer to added PPh(3), the kinetics of which has been studied with double mixing experiments in a cryogenic stopped-flow apparatus. The results show that the decay of the THF intermediate is kinetically controlled by the dissociation of a THF ligand, a conclusion supported by the observation of saturation kinetic behavior with respect to PPh(3), inhibition by added THF, and invariant saturation rate constants for the oxidation of various phosphines. It is proposed that the proximity of the reducing substrate to the peroxide ligand on the diiron coordination sphere facilitates the oxygen-atom transfer. This unique investigation of the reaction of an O(2) adduct of a biomimetic tetracarboxylatodiiron(II,II) complex provides a synthetic precedent for understanding the electrophilic reactivity of like adducts in the active sties of nonheme diiron enzymes.

Copper(I)-phenolate complexes as models of the reduced active site of galactose oxidase: synthesis, characterization, and O2 reactivity.

The Cu(I)-phenolate complexes (1)LCu and (2)LCu and the Cu(I)-phenol complex [H(2)LCu(CNC(6)H(3)Me(2))]BArF(4) were prepared and structurally characterized by X-ray crystallography, where (1)L(-) and (2)L(-) are ligands comprised of a 2,4-di- tert-butylphenolate linked to 1-isopropyl-1,5-diazacyclooctane or 1,4-diisopropyl-1,4,7-triazacyclononane, respectively. The reduced galactose oxidase (GAO) structural models (1)LCu and (2)LCu were found to be highly reactive with O(2), and through combined stopped-flow kinetic and EPR, UV-vis, and resonance Raman spectroscopic studies of the oxygenation of (2)LCu at low temperature, new intermediates relevant to those postulated for the active site oxidation step of the GAO catalytic cycle were identified. The oxygenation was shown by kinetics experiments to proceed via initial binding of O(2) to yield a green, unusually thermodynamically stable 1:1 adduct, (2)LCu(O(2)). Symmetric (eta(2)) binding of a superoxo ligand was indicated by oxygen-isotope-sensitive features in resonance Raman spectra obtained in batch experiments; peaks at nu((16)O(2))=1120 cm(-1), nu((18)O(16)O)=1093 cm(-1), and nu((18)O(2))=1058 cm(-1) were assigned as O-O stretching vibrations. These data represent the first experimental evidence for such superoxide coordination in complexes of tetradentate tripodal ligands and provide new precedent for how O(2) may bind at the reduced GAO active site. The 1:1 Cu/O(2) adduct subsequently evolves into a metastable purple species that is only observable under conditions of substoichiometric O(2). The kinetics of formation of this transient species are second order overall (rate= k'(2)[(2)LCu(O(2))][(2)LCu]). It exhibits an absorption band with lambda(max)=565 nm (epsilon=17900 M(-1) cm(-1)) and multiple oxygen-isotope-sensitive nu(Cu-O) and nu(O-O) features in the respective regions 500-550 cm(-1) and 700-850 cm(-1) in Raman spectra, with excitation-wavelength-dependent intensities that correlate with the 565 nm absorption feature. On the basis of the combined data available, the presence of multiple isomeric peroxodicopper species in the transient purple solution is postulated.

Copper chemistry of beta-diketiminate ligands: monomer/dimer equilibria and a new class of bis(mu-oxo)dicopper compounds.

A series of Cu(I) and Cu(II) complexes of a variety of beta-diketiminate ligands (L(-)) with a range of substitution patterns were prepared and characterized by spectroscopic, electrochemical, and, in several cases, X-ray crystallographic methods. Specifically, complexes of the general formula [LCuCl](2) were structurally characterized and their magnetic properties assessed through EPR spectroscopy of solutions and, in one instance, by variable-temperature SQUID magnetization measurements on a powder sample. UV-vis spectra indicated reversible dissociation to 3-coordinate monomers LCuCl in solution at temperatures above -55 degrees C. The Cu(I) complexes LCu(MeCN) exhibited reversible Cu(I)/Cu(II) redox couples with E(1/2) values between +300 and +520 mV versus NHE (cyclic voltammetry, MeCN solutions). These complexes were highly reactive with O(2), yielding intermediates that were identified as rare examples of neutral bis(mu-oxo)dicopper complexes on the basis of their EPR silence, diagnostic UV-vis absorption data, and O-isotope-sensitive resonance Raman spectroscopic features. The structural features of the compounds [LCuCl](2) and LCu(MeCN) as well as the proclivity to form bis(mu-oxo)dicopper products upon oxygenation of the Cu(I) complexes are compared to data previously reported for complexes of more sterically hindered beta-diketiminate ligands (Aboelella, N. W.; Lewis, E. A.; Reynolds, A. M.; Brennessel, W. W.; Cramer, C. J.; Tolman, W. B. J. Am. Chem. Soc. 2002, 124, 10600. Spencer, D. J. E.; Aboelella, N. W.; Reynolds, A. M.; Holland, P. L.; Tolman, W. B. J. Am. Chem. Soc. 2002, 124, 2108. Holland, P. L.; Tolman, W. B. J. Am. Chem. Soc. 1999, 121, 7270). The observed structural and reactivity differences are rationalized by considering the steric influences of both the substituents on the flanking aromatic rings and those present on the beta-diketiminate backbone.

Snapshots of dioxygen activation by copper: the structure of a 1:1 Cu/O(2) adduct and its use in syntheses of asymmetric Bis(mu-oxo) complexes.

The X-ray structure of a 1:1 Cu/O(2) adduct revealed side-on (eta(2)) O(2) coordination. Density functional calculations corroborated the structure, indicated a significant contribution of a Cu(III)-(O(2)(2-)) resonance form, and provided insights into the key bonding interactions. Reaction of a 1:1 adduct supported by a slightly different beta-diketiminate ligand with Cu(I) reagents resulted in the formation of novel asymmetric bis(mu-oxo) complexes that were identified by EPR, UV-vis, and Raman spectroscopy, as well as by an X-ray structure in one instance.

beta-diketiminate ligand backbone structural effects on Cu(I)/O(2) reactivity: unique copper-superoxo and bis(mu-oxo) complexes.

Copper(I) and -(II) complexes of beta-diketiminate ligands with identical flanking 2,6-diisopropylphenyl groups but divergent backbone substitution patterns were prepared and structurally characterized, and reactions of the Cu(I) species with O(2) at low temperature were explored. Despite being far removed from the coordinated metal ion, the different backbone patterns significantly influence the steric encumbrance exerted by the ligands, as revealed by differences in (a) the structural features of the Cu(I) and Cu(II) complexes and (b) the course of the oxygenation reactions of the Cu(I) compounds. With the less hindered ligand, a rare example of a neutral bis(mu-oxo)dicopper complex was identified on the basis of its diagnostic spectral features (UV-vis, resonance Raman, EPR) and the stoichiometry of O(2) uptake (Cu:O(2) = 2:1). In contrast, oxygenation of the Cu(I) complexes supported by the more hindered ligands yielded novel (superoxo)copper complexes, identified by a Cu:O(2) ratio of 1:1, a lack of an EPR signal, and O-isotope sensitive resonance Raman spectral features (nu(O)(-)(O) = 968 cm(-1), Delta(18)O(2) = 51 cm(-1)). Symmetric coordination of the superoxo ligand is proposed on the basis of Raman data acquired using (16)O(18)O (single peak at 943 cm(-1)).